Project summary The goal of this proposal is to advance in vitro modeling of human heart disease using genome-edited and patient-derived iPSCs, to use these models to gain new insights into disease pathogenesis, and to develop new therapeutic strategies. We focus on three monogenic cardiac diseases, Barth syndrome (BTHS), cate- cholaminergic polymorphic ventricular tachycardia (CPVT), and arrhythmogenic cardiomyopathy (ACM; also known as arrhythmogenic right ventricular cardiomyopathy/dysplasia). These disorders represent major classes of inherited heart disease, namely disorders of cardiac rhythm (CPVT; ACM) and contraction (BTHS; ACM). No targeted therapies are available for these disorders, and current management options are far from ideal, resulting in tragic deaths or cardiac transplantation. Our studies of these diseases will push the envelope of in vitro disease models in four principle ways: (1) by refining in vitro systems to better reflect the physiology of native myocardium; (2) by objectively evaluating the ability of induced pluripotent stem cell-derived cardiomyocyte (iPSC-CM) models to capture inter-individual variation between patients; (3) by identifying novel therapies through either improved mechanistic understanding or unbiased screening; and (4) by performing proof-of-concept ?Clinical trials in a dish?, in which responses of engineered cell and tissue models are compared to responses of mammalian models or patients. In the UG3 Phase, we will develop physiological assay systems of these three monogenic cardiac diseases (Aim 1). These assay systems will scale from cell pairs to three dimensional engineered ventricles, providing the range of systems necessary to address challenges spanning high throughput screening to disease pathogenesis to in vitro ?clinical trials?. In the UH3 Phase, we will use the 2D tissues and 3D ventricles to discover novel treatments through screens and mechanistic studies (Aim 2). We will use the 3D ventricles to perform ?Clinical trials in a dish? (Aim 3), to determine the extent to which iPSC-based models capture inter-individual variation and to measure the therapeutic responses of a panel of patient microphysiological models. Our vertically integrated, multidisciplinary approach will bring together cardiac biologists, bioengineers, bioinformaticians, and clinicians to advance the state of the art for in vitro cardiac disease modeling. The impact will extend well beyond the three rare diseases directly studied by improving cardiac disease models and providing data on the usefulness of iPSC-CMs for capturing individual patient phenotypes. Creation of in vitro models of normal human organs would also greatly expedite drug development, by increasing the precision and speed of drug safety testing. Our deliverables include advances in iPSC-CM differentiation; novel bioengineered systems to assay iPSC-CM physiological properties; new insights into the pathogenesis of three representative cardiac diseases; and identification of therapeutic targets and lead compounds for disease treatment.